Identification of Transformation Products for Benzotriazole, Triclosan, and Trimethoprim by Aerobic and Anoxic-Activated Sludge

Abstract

This study identified biotransformation products formed from three anthropogenic trace organic compounds (TOrCs) (benzotriazole, triclosan, and trimethoprim) in aerobic and anoxic sludge from a biological nutrient removal (BNR) wastewater treatment system. Liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QTOF) was used to identify intermediate transformation products. A user-defined compound library was used to target unknowns in samples with a mix of TOrCs without requiring chemical standards and developed using EAWAG-BDD predictive software, which included 37 potential transformation products. Biotransformation batch experiments were conducted using activated sludge from anoxic and aerobic redox regimes of a BNR treatment plant located in Southern Nevada, United States. Four intermediates were observed for benzotriazole in both aerobic and anoxic activated sludge, which consisted of two isomers of hydroxy benzotriazole and two isomers of methoxy benzotriazole. Four intermediates were observed for trimethoprim under aerobic conditions, which formed 2,4-diaminopyrimidin-5-yl)(3,4,5-trimethoxyphenyl)methanol (TMP 306) and 2,6-diamino-5-hydroxy-5-(3,4,5-trimethoxybenzyl)-5,6-dihydropyrimidin-4(1H)-one (TMP 324) and two demethylation isomers of desmethyl trimethoprim. Triclosan had one confirmed transformation product, triclosan-o-sulfate, formed in both conditions. The identification of these transformation products will allow for more thorough risk assessments to be performed for the target TOrCs.

A Review on the Fate of Legacy and Alternative Antimicrobials and Their Metabolites during Wastewater and Sludge Treatment

“…Chen et al [57] observed that TCS was transformed into TCS-SO 4 through sulfation during a batch simulated aerobic activated sludge process. This agrees with a later batch study by Trenholm et al [84], who observed the rapid formation of TCS-SO 4 under aerobic conditions and slower formation of TCS-SO 4 under anoxic conditions using biological nutrient removal (BNR) sludge. Although Chen et al [57] did not observe any decrease in TCS-SO 4 once it was formed over 10 days, Trenholm et al [84] observed that TCS-SO 4 levels began to decrease after 5 days under aerobic conditions and observed the formation of a secondary TCS-SO 4 metabolite, whose mass matched 3,5-dichloro-2-hydroxy-benzene sulfonate (or a closely related isomer thereof).…”

Pharmaceutical Biotransformation is Influenced by Photosynthesis and Microbial Nitrogen Cycling in a Benthic Wetland Biomat

“…In terms of transformation reactions, we observed oxidation of the hydroxyl moiety to carboxy-emtricitabine in the case of emtricitabine (Figure S13b), and trimethoprim has previously been shown to undergo biotransformation via hydroxylation or demethylation during biological wastewater treatment. 118,119 These transformations are not consistent with the expected reactive sites for copper-containing enzymes encoded by genes in our metagenomes from the field (e.g., amine oxidases or laccases), which would more likely result in oxidation of the pyrimidine 2,4-diamine group in trimethoprim, 120 the product of which has not been observed under wastewater treatment conditions. Collectively, these data provide supporting evidence that our allylthiourea biotransformation inferences were not impacted by copper-dependent enzymes outside of AMO.…”

“…However, inspection of MAGs associated with ASVs significantly suppressed by allylthiourea (Figure b, Table S4; p adj < 0.005) revealed genes encoding for copper-dependent amine oxidases (i.e., Vicinamibacterales, Pirellulaceae, Pirellula_B gen), and several scaffolds belonging to other MAGs or that were unbinned also encoded for both copper-dependent amine oxidases and laccases, which have the potential to co-metabolically biotransform amine and (poly)­phenol containing compounds. In terms of transformation reactions, we observed oxidation of the hydroxyl moiety to carboxy-emtricitabine in the case of emtricitabine (Figure S13b), and trimethoprim has previously been shown to undergo biotransformation via hydroxylation or demethylation during biological wastewater treatment. , These transformations are not consistent with the expected reactive sites for copper-containing enzymes encoded by genes in our metagenomes from the field (e.g., amine oxidases or laccases), which would more likely result in oxidation of the pyrimidine 2,4-diamine group in trimethoprim, the product of which has not been observed under wastewater treatment conditions. Collectively, these data provide supporting evidence that our allylthiourea biotransformation inferences were not impacted by copper-dependent enzymes outside of AMO.…”

Pharmaceutical Attenuation Differs within Woodchip-Based Lignocellulose Bioreactors across Nitrate- and Sulfate-Reducing Conditions

“…As EBCT increased, the percent removal of trimethoprim in the sulfate-reducing columns (0 and 10 mg/L-N) also increased, with the largest difference observed in the 0 mg/L-N and 10 mg/L-N columns (Figure B and Table S2). Trimethoprim removal was accompanied by the production of low concentrations of desmethyl trimethoprim, a biotransformation product of trimethoprim. , Higher concentrations of this product were seen in the effluent of the sulfate-reducing columns (0.13 ± 0.10 μg/L) than in the nitrate-reducing columns (0.03 ± 0.01 μg/L). Unfortunately, desmethyl trimethoprim was only quantified in samples collected during the last two time points of the experiment, limiting opportunities for statistical analysis.…”